The invention relates to a vaccine for hemorrhagic fever with renal syndrome caused by hantavirus infections.
The Hantavirus genus of the family Bunyaviridae includes a number of rodent-borne viruses that can cause hemorrhagic fever with renal syndrome (HFRS) or hantavirus pulmonary syndrome (HPS). At least four hantaviruses cause HFRS: Hantaan (HTNV), Seoul (SEOV), Dobrava (DOBV), and Puumala (PUUV) viruses. HFRS presents with sudden fever, chills, nausea, headache, and backache. Early symptoms of severe HFRS often also include facial flushing, conjunctivitis, and petechial rash. Death can occur due to vascular leakage leading to low blood pressure, acute shock, and renal failure. There are no FDA-licensed vaccines for HFRS, but an inactivated, rodent-brain-derived HTNV vaccine is commercially available in Korea, and several inactivated cell culture-derived HTNV and SEOV vaccines have been developed in China [1,2].
Despite the use of these vaccines for more than a decade, HFRS remains a significant public health threat in Asia with thousands of hospitalized cases reported each year in China [3-5]. Several hundred to thousands of HFRS cases due to PUUV or DOBV infections are reported each year in Europe, Scandinavia, and Russia, with the greatest incidences observed in Finland (25,000 cases from 1979 to 2006) and western Russia (˜89,000 cases from 1996 [6]. Inactivated vaccines have not been developed in Europe, in part because PUUV is difficult to grow in cell culture to high enough titers for scale-up, and rodent brain-derived vaccines are not considered desirable. Moreover, because DOBV and PUUV both cause HFRS in the same geographic region, and because there is little or no cross-protective immunity between PUUV and the other HFRS-causing hantaviruses [7,8], a comprehensive vaccine for European HFRS will need to elicit protective immunity to both viruses.
To date, two recombinant DNA vaccines for HFRS have been tested in early clinical studies. The first tested was a vaccinia virus (VACV)-vectored vaccine, developed and evaluated in Phases 1 and 2 clinical studies by USAMRIID [9,10]. The vaccine expressed two of the three gene segments of HTNV: the M segment, which encodes the envelope glycoproteins (Gn and Gc), and the S segment, which encodes the nucleocapsid protein (N). In general, animal studies have shown that neutralizing antibodies to Gn and Gc are the best measurable correlate of protective immunity [8, 11-13]. This earlier study found that the recombinant VACV vaccine elicited neutralizing antibodies against HTNV in VACV-naïve individuals, but was poorly immunogenic in VACV-immune volunteers [9]. Consequently, the vaccine developers changed strategies to a DNA vaccine platform, which was not adversely affected by preexisting vector immunity and which offered additional flexibility for producing combination vaccines. In addition to flexibility, DNA is an attractive vaccine platform in terms of ease of engineering and manufacturing as well as safety.
USAMRIID investigators have so far conducted two Phase 1 clinical studies with DNA vaccines for HFRS using DNA derived from HTNV and from PUUV M segments. The two-part DNA vaccine strategy was used because vaccination with the HTNV M gene-based DNA vaccine protects animals from infection with HTNV, DOBV and SEOV, but not from PUUV infection. PUUV M gene-based DNA vaccine protects against infection with PUUV [7-8].
The first two clinical studies of the HTNV and PUUV DNA vaccines were performed using a PUUV M segment vaccine that was genetically optimized (US 2010/0323024A1, incorporated herein by reference in its entirety). The HTNV component, however, was not optimized, because unlike the PUUV DNA, which required optimization for gene expression, the HTNV DNA construct showed strong gene expression without optimization. It could not be anticipated, therefore, that a similar optimization was either necessary or would offer a benefit over the non-optimized DNA for immunogenicity. Further and formerly, an extraneous gene sequence was required for the expression of the non-optimized HTNV gene, U.S. Pat. No. 7,217,812, incorporated by reference, herein, in its entirety.
In the first clinical study of the DNA vaccines, HTNV and PUUV M segments were delivered by particle mediated epidermal delivery (PMED). The advantage of intraepidermal delivery of the vaccine is twofold. The DNA is easily taken up by cells at the site of delivery or by cells in the draining lymph nodes where the antigen encoded by those cells is reprocessed by specialized antigen-presenting cells to elicit an immune response, and this approach uses 1000-fold less DNA than needle administration.
The vaccines were given as separate administrations because of results from animal studies, which showed that if the HTNV vaccine is mixed with the PUUV vaccine, then only neutralizing antibodies to PUUV are elicited [25]. This finding was not expected, because it was possible to obtain strong responses to the individual vaccines or to both vaccines when they were delivered simultaneously, but as separate inoculations, to a single animal. In addition, it was not possible to overcome this interference by adjusting the ratio of HTNV: PUUV DNA even as high as 10:1 (FIG. 8B). Other attempts to produce modified constructs that were chimeras of both the HTNV and PUUV genes also failed to elicit antibody responses to both HTNV and PUUV (unpublished information). The outcome of the interference study is summarized in Example 1.
In a second Phase 1 clinical study of the same two DNA vaccines, the DNAs were given separately or as a mixture by intramuscular electroporation (IM-EP). With this delivery method, the vaccines are injected into muscles and a rapid electrical pulse is applied to facilitate uptake of the DNA into the muscle cells. Because a larger number of host cells receive the vaccines than when they are delivered by PMED, it was anticipated that there might be some response to both vaccines. As expected, however, interference was still a problem in individuals receiving the mixed vaccines, with better responses obtained to the PUUV vaccine than to the HTNV vaccine as shown in Example 2.
Delivery of the vaccine can also be by nanoparticle encapsulation of the vaccine via various methods, including aerosol delivery of the nanoparticles.
The present invention provides a combination vaccine to protect against HFRS. The invention consists of an optimized HTNV M segment vaccine, which solves the problem of interference in the bivalent vaccine. Unlike the non-optimized HTNV vaccine used in previous studies, the vaccine of the invention can be mixed with a similarly optimized PUUV-based vaccine to elicit neutralizing antibodies against both viruses. The invention provides a safe, economical, flexible and effective vaccine for the protection of humans from HFRS caused by infection with HTNV, SEOV, PUUV and/or DOBV.